The goal of this proposal is to develop a high throughput screening assay to identify small molecules that enhance plasma membrane repair in muscle cells. Membrane repair after injury is a critical process that entails the formation, translocation and fusion of a membrane patch with the disrupted plasma membrane. Mutations that knockout the muscle membrane protein, dysferlin, cause limb girdle muscular dystrophy type 2B (LGMD2B) and a distal dystrophy known as Miyoshi myopathy (MM). We have shown that muscle membrane repair is impaired in myotubes lacking dysferlin. The premise of this proposal is that molecules that improve the membrane repair response in muscle cells will be beneficial in patients with Dysferlin deficiency and possibly other muscle diseases. Our specific aims are to: (1) Optimize our previously validated, quantitative membrane injury assay in a 96-well format; (2) Perform a high-throughput screen to identify small molecules that augment the membrane repair response in normal (dysferlin-positive) and dystrophic (dysferlin- deficient) muscle cell lines; (3) Validate the compounds that provide "hits" from each screen using biochemical and functional secondary assays. This proposal is significant for several reasons: (1) To our knowledge, it is the first to screen for molecules that enhance membrane repair; (2) Small molecules found using this platform (Aim 2a and 2b) my be beneficial not only in patients with impaired membrane repair systems, but also in any subjects suffering from exercise-induced muscle injury; (3) This platform can also be used to test the efficacy of other therapeutics in dysferlin-deficiency e.g. gene therapy and cell therapy. The rationale behind this approach is: (1) We know that the physical disruption of muscle membrane induces an injury that results in a patch fusion repair response and that this response is quantifiably retarded in dysferlin-deficient cells; (2) A high-throughput screen is the best possible method for finding molecules that might alter repair and such molecules could have beneficial effects in both normal and dystrophic muscle.